The primary goal of this continuing project is to describe the metabolism of chondroitin SO4 in chick embryo tissues. In the coming year the work will deal with both the biosynthetic steps and the catobolism. These processes will be studied using monolayer cultures of chondrocytes and cell-free extracts prepared from embryonic liver and cartilage. In the experiments using monolayers of chondrocytes, the effects of culture conditions on the rate of chondroitin SO4 synthesis and secretion will be studied using 35SO24- as a labeled precursor. The biosynthetic products will be characterized according to molecular size, protein content, and ratio of 6-sulfated to 4-sulfated galactosamine residues. As noted, the studies of enzyme activity in cell-free extracts will deal with both biosynthetic and degradative steps. The catabolic enzymes to be studied include hexosaminidase and the 4- and 6-sulfatases. Assays for the action of these enzymes on oligosaccharides prepared from chondroitin SO4's and hyaluronic acid have been developed, and these will be applied in a determination of the kinetic properties and the substrate specificities of these enzymes. These same oligosaccharides will be used as acceptors in studies of sulfuryltransferase and glucuronosyltransferase reactions. Once again the kinetic properties and the oligosaccharide specificities of these enzymes will be determined, and the products will be characterized.